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RRM2 and CDC6 are novel effectors of XBP1-mediated endocrine resistance and predictive markers of tamoxifen sensitivity
BACKGROUND: Endocrine-resistant breast cancers have elevated expression of XBP1, where it drives endocrine resistance by controlling the expression of its target genes. Despite the in-depth understanding of the biological functions of XBP1 in ER-positive breast cancer, effectors of endocrine resista...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10061897/ https://www.ncbi.nlm.nih.gov/pubmed/36997866 http://dx.doi.org/10.1186/s12885-023-10745-1 |
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author | Barua, David Sultana, Afrin Islam, Md Nahidul Cox, Fergus Gupta, Ananya Gupta, Sanjeev |
author_facet | Barua, David Sultana, Afrin Islam, Md Nahidul Cox, Fergus Gupta, Ananya Gupta, Sanjeev |
author_sort | Barua, David |
collection | PubMed |
description | BACKGROUND: Endocrine-resistant breast cancers have elevated expression of XBP1, where it drives endocrine resistance by controlling the expression of its target genes. Despite the in-depth understanding of the biological functions of XBP1 in ER-positive breast cancer, effectors of endocrine resistance downstream of XBP1 are poorly understood. The aim of this study was to identify the XBP1-regulated genes contributing to endocrine resistance in breast cancer. METHODS: XBP1 deficient sub-clones in MCF7 cells were generated using the CRISPR-Cas9 gene knockout strategy and were validated using western blot and RT-PCR. Cell viability and cell proliferation were evaluated using the MTS assay and colony formation assay, respectively. Cell death and cell cycle analysis were determined using flow cytometry. Transcriptomic data was analysed to identify XBP1-regulated targets and differential expression of target genes was evaluated using western blot and qRT-PCR. Lentivirus and retrovirus transfection were used to generate RRM2 and CDC6 overexpressing clones, respectively. The prognostic value of the XBP1-gene signature was analysed using Kaplan–Meier survival analysis. RESULTS: Deletion of XBP1 compromised the upregulation of UPR-target genes during conditions of endoplasmic reticulum (EnR) stress and sensitized cells to EnR stress-induced cell death. Loss of XBP1 in MCF7 cells decreased cell growth, attenuated the induction of estrogen-responsive genes and sensitized them to anti-estrogen agents. The expression of cell cycle associated genes RRM2, CDC6, and TOP2A was significantly reduced upon XBP1 deletion/inhibition in several ER-positive breast cancer cells. Expression of RRM2, CDC6, and TOP2A was increased upon estrogen stimulation and in cells harbouring point-mutants (Y537S, D538G) of ESR1 in steroid free conditions. Ectopic expression of RRM2 and CDC6 increased cell growth and reversed the hypersensitivity of XBP1 KO cells towards tamoxifen conferring endocrine resistance. Importantly, increased expression of XBP1-gene signature was associated with poor outcome and reduced efficacy of tamoxifen treatment in ER-positive breast cancer. CONCLUSIONS: Our results suggest that RRM2 and CDC6 downstream of XBP1 contribute to endocrine resistance in ER-positive breast cancer. XBP1-gene signature is associated with poor outcome and response to tamoxifen in ER-positive breast cancer. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12885-023-10745-1. |
format | Online Article Text |
id | pubmed-10061897 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-100618972023-03-31 RRM2 and CDC6 are novel effectors of XBP1-mediated endocrine resistance and predictive markers of tamoxifen sensitivity Barua, David Sultana, Afrin Islam, Md Nahidul Cox, Fergus Gupta, Ananya Gupta, Sanjeev BMC Cancer Research BACKGROUND: Endocrine-resistant breast cancers have elevated expression of XBP1, where it drives endocrine resistance by controlling the expression of its target genes. Despite the in-depth understanding of the biological functions of XBP1 in ER-positive breast cancer, effectors of endocrine resistance downstream of XBP1 are poorly understood. The aim of this study was to identify the XBP1-regulated genes contributing to endocrine resistance in breast cancer. METHODS: XBP1 deficient sub-clones in MCF7 cells were generated using the CRISPR-Cas9 gene knockout strategy and were validated using western blot and RT-PCR. Cell viability and cell proliferation were evaluated using the MTS assay and colony formation assay, respectively. Cell death and cell cycle analysis were determined using flow cytometry. Transcriptomic data was analysed to identify XBP1-regulated targets and differential expression of target genes was evaluated using western blot and qRT-PCR. Lentivirus and retrovirus transfection were used to generate RRM2 and CDC6 overexpressing clones, respectively. The prognostic value of the XBP1-gene signature was analysed using Kaplan–Meier survival analysis. RESULTS: Deletion of XBP1 compromised the upregulation of UPR-target genes during conditions of endoplasmic reticulum (EnR) stress and sensitized cells to EnR stress-induced cell death. Loss of XBP1 in MCF7 cells decreased cell growth, attenuated the induction of estrogen-responsive genes and sensitized them to anti-estrogen agents. The expression of cell cycle associated genes RRM2, CDC6, and TOP2A was significantly reduced upon XBP1 deletion/inhibition in several ER-positive breast cancer cells. Expression of RRM2, CDC6, and TOP2A was increased upon estrogen stimulation and in cells harbouring point-mutants (Y537S, D538G) of ESR1 in steroid free conditions. Ectopic expression of RRM2 and CDC6 increased cell growth and reversed the hypersensitivity of XBP1 KO cells towards tamoxifen conferring endocrine resistance. Importantly, increased expression of XBP1-gene signature was associated with poor outcome and reduced efficacy of tamoxifen treatment in ER-positive breast cancer. CONCLUSIONS: Our results suggest that RRM2 and CDC6 downstream of XBP1 contribute to endocrine resistance in ER-positive breast cancer. XBP1-gene signature is associated with poor outcome and response to tamoxifen in ER-positive breast cancer. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12885-023-10745-1. BioMed Central 2023-03-30 /pmc/articles/PMC10061897/ /pubmed/36997866 http://dx.doi.org/10.1186/s12885-023-10745-1 Text en © The Author(s) 2023 https://creativecommons.org/licenses/by/4.0/Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) . The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/ (https://creativecommons.org/publicdomain/zero/1.0/) ) applies to the data made available in this article, unless otherwise stated in a credit line to the data. |
spellingShingle | Research Barua, David Sultana, Afrin Islam, Md Nahidul Cox, Fergus Gupta, Ananya Gupta, Sanjeev RRM2 and CDC6 are novel effectors of XBP1-mediated endocrine resistance and predictive markers of tamoxifen sensitivity |
title | RRM2 and CDC6 are novel effectors of XBP1-mediated endocrine resistance and predictive markers of tamoxifen sensitivity |
title_full | RRM2 and CDC6 are novel effectors of XBP1-mediated endocrine resistance and predictive markers of tamoxifen sensitivity |
title_fullStr | RRM2 and CDC6 are novel effectors of XBP1-mediated endocrine resistance and predictive markers of tamoxifen sensitivity |
title_full_unstemmed | RRM2 and CDC6 are novel effectors of XBP1-mediated endocrine resistance and predictive markers of tamoxifen sensitivity |
title_short | RRM2 and CDC6 are novel effectors of XBP1-mediated endocrine resistance and predictive markers of tamoxifen sensitivity |
title_sort | rrm2 and cdc6 are novel effectors of xbp1-mediated endocrine resistance and predictive markers of tamoxifen sensitivity |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10061897/ https://www.ncbi.nlm.nih.gov/pubmed/36997866 http://dx.doi.org/10.1186/s12885-023-10745-1 |
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